Manufacture of Degarelix

ABSTRACT

The present invention provides methods for producing a lyophilized degarelix product which, upon reconstitution with water for injection in an amount of 20 mg/ml, shows a viscosity of up to 15 mPas. The present invention also provides a lyophilized degarelix drug substance which shows, upon dissolution in water in an amount of 20 mg/ml, a viscosity of up to 3.2 mPas, and processes for providing this lyophilized degarelix drug substance.

This is a continuation of application Ser. No. 15/420,156, filed Jan.31, 2017 (now U.S. Pat. No. 10,172,906 B2, issued Jan. 8, 2019), whichis a continuation of application Ser. No. 14/403,775, filed Nov. 25,2014 (now U.S. Pat. No. 9,592,266 B2, issued on Mar. 14, 2017), which isa national stage application under 35 U.S.C. § 371 of InternationalApplication No. PCT/EP2013/061264, filed May 31, 2013, which claimspriority of European Patent Application No. 12170454, filed Jun. 1,2012; the contents of each application are incorporated herein byreference.

TECHNICAL FIELD

The present invention relates to a manufacturing process for preparingDegarelix.

BACKGROUND OF THE INVENTION

Prostate cancer is a leading cause of morbidity and mortality for men inthe industrialised world. Degarelix, also known as FE200486, is a thirdgeneration gonadotropin releasing hormone (GnRH) receptor antagonist (aGnRH blocker) that has been developed and approved for prostate cancerpatients in need of androgen ablation therapy (Doehn et al., Drugs 2006,vol. 9, No. 8, pp. 565-571; WO 09846634). Degarelix acts by immediateand competitive blockade of GnRH receptors in the pituitary and, likeother GnRH antagonists, does not cause an initial stimulation ofluteinizing hormone production via the hypothalamic-pituitary-gonadalaxis, and therefore does not cause testosterone surge or clinical flare(Van Poppel, Cancer Management and Research, 2010:2 39-52; Van Poppel etal., Urology, 2008, 71(6), 1001-1006); James, E. F. et al., Drugs, 2009,69(14), 1967-1976).

Degarelix is a synthetic linear decapeptide containing seven unnaturalamino acids, five of which are D-amino acids. It has ten chiral centersin the back bone of the decapeptide. The amino acid residue at position5 in the sequence has an additional chiral center in the side-chainsubstitution giving eleven chiral centers in total. Its CAS registrynumber is 214766-78-6 (of free base) and it is commercially availableunder the Trademark Firmagon™. The drug substance is chemicallydesignated as D-Alaninamide,N-acetyl-3-(2-naphthalenyl)-D-alanyl-4-chloro-D-phenylalanyl-3-(3-pyridinyl)-D-alanyl-L-seryl-4-[[[(4S)-hexahydro-2,6-dioxo-4-pyrimidinyl]carbonyl]amino]-L-phenylalanyl-4-[(aminocarbonyl)amino]-D-phenylalanyl-L-leucyl-N6-(1-methylethyl)-L-lysyl-L-prolyl-and is represented by the chemical structure below (in the followingalso referred to as Formula I):

The structure of Degarelix can also be represented as:Ac-D-2Nal-D-4Cpa-D-3Pal-Ser-4Aph(L-Hor)-D-4Aph(Cbm)-Leu-Lys(iPr)-Pro-D-Ala-NH₂where Ac is acetyl, 2Nal is 2-naphthylalanine, 4Cpa is4-chlorophenylalanine, 3Pal is 3-pyridylalanine, Ser is serine, 4Aph is4-aminophenylalanine, Hor is hydroorotyl, Cbm is carbamoyl, Leu isleucine, Lys(iPr) is N6-isopropyllysine, Pro is proline and Ala isalanine.

For the purposes of describing this invention, each amino acid inDegarelix will be given the shorthand notation as follows:

AA₁ is D-2Nal, AA₂ is D-4Cpa, AA₃ is D-3Pal, AA₄ is Ser, AA₅ is4Aph(L-Hor), AA₆ is D-Aph(Cbm), AA₇ is Leu, AA₈ is Lys(iPr), AA₉ is Proand AA₁₀ is D-Ala.

Thus, Degarelix can be represented as Ac-AA₁-AA₁₀-NH₂.

Degarelix has previously been prepared using Boc-solid phase peptidesynthesis (SPPS) methodology as reported in WO 98/46634 and Jiang etal., J. Med. Chem. 2001, 44, 453-467.

WO2010/12835 and WO2011/066386 describe the preparation of degarelixusing an Fmoc strategy. WO2012/055905 and WO2012/055903 describe liquidphase syntheses of degarelix.

SUMMARY OF THE INVENTION

The physicochemical characterization of degarelix has shown that thisdecapeptide has the ability to self-associate, and eventually form gelsin aqueous solution. Self-aggregation makes this compound build-up adepot in situ when injected subcutaneously or intramuscularly. Thedegarelix depot was shown to provide a sustained release of the activeover months depending on the dosage. At present, the drug isadministered in dosages of 120 mg (40 mg/ml) for first injection, and of80 mg (20 mg/ml) for sustained release over one month.

The present inventors have surprisingly found that viscosity, and hencesustained release properties and bioavailability, of the reconstituteddrug product can be controlled through processing of the crude peptide(e.g. obtained by Fmoc strategy, liquid phase synthesis or anotherroute) into the drug substance. The viscosity associated with the drugsubstance surprisingly correlates with the viscosity associated with thedrug product, even after a further reconstitution and lyophilisation.The viscosity of the drug product has to be controlled within a range ofup to 15 mPas, preferably within a range of 2 to 12 mPas, to obtain thedesired depot formation and thus sustained release. The presentinvention provides processes that allow the manufacture of drug productsthat show this viscosity, as determined upon reconstitution with thereconstitution fluid at a concentration of 20 mg degarelix per mlreconstitution fluid.

In a first aspect, the present invention thus provides a method forcontrolling the viscosity of a degarelix product to be no greater than15 mPas, preferably within a range of 2 to 12 mPas, as determined uponreconstitution with water for injection in an amount of 20 mg degarelixfree base/ml, comprising the steps of:

-   -   1. Providing a lyophilized degarelix drug substance which shows        a viscosity of up to 3.2 mPas, as determined upon dissolution in        water containing mannitol (2.5% w/V) in an amount of 20 mg        degarelix free base/ml    -   2. Dissolving the lyophilized degarelix drug substance in        mannitol-containing water to provide an aqueous        degarelix-mannitol mixture;    -   3. Lyophilizing the aqueous degarelix-mannitol mixture to        provide the degarelix drug product.

In a second aspect, the present invention provides a method forproducing a lyophilized degarelix product which shows a viscosity of upto 15 mPas, preferably within a range of 2 to 12 mPas, as determinedupon reconstitution with water for injection in an amount of 20 mgdegarelix free base/ml, comprising the steps of:

-   -   1. Providing a lyophilized degarelix drug substance which shows        a viscosity of up to 3.2 mPas, as determined upon dissolution in        water containing mannitol (2.5% w/V) in an amount of 20 mg        degarelix free base/ml    -   2. Dissolving the lyophilized degarelix drug substance in        mannitol-containing water to provide an aqueous        degarelix-mannitol mixture; and    -   3. Lyophilizing the aqueous degarelix-mannitol mixture to        provide the degarelix drug product.

In a third aspect, the present invention provides a method for producingdegarelix drug product comprising a lyophilized degarelix drug productand a liquid for reconstitution (reconstitution fluid) which, uponreconstitution with said liquid in an amount of 20 mg degarelix freebase/ml, shows a viscosity of up to 15 mPas, preferably within a rangeof 2 to 12 mPas, comprising the steps of:

-   -   1. Dissolving a lyophilized degarelix drug substance in a        mannitol containing aqueous solution to provide a        degarelix-mannitol mixture;    -   2. Lyophilizing the degarelix-mannitol mixture to provide the        degarelix drug product, wherein a viscosity-reducing agent is        added to the degarelix-mannitol mixture prior to lyophilization.

The present invention also provides a lyophilized degarelix drugsubstance which shows, upon dissolution in water containing 2.5 wt. %mannitol in an amount of 20 mg degarelix free base/ml, a viscosity of upto 3.2 mPas, and processes for providing this lyophilized degarelix drugsubstance.

Moreover, the present invention provides a degarelix drug productcomprising a lyophilized degarelix drug product and a reconstitutionfluid which, upon reconstitution with said reconstitution fluid in anamount of 20 mg degarelix free base/ml, shows a viscosity of up to 15mPas, preferably within a range of 2 to 12 mPas, and contains aviscosity-reducing agent in an amount of 0.001 to 5 mg/ml.

FIGURES

FIG. 1 shows the relationship between drug substance viscosity and drugproduct viscosity.

FIGS. 2 and 3 show the relationship between dynamic viscosity ofdegarelix dosing suspension (20 mg/ml) and degarelix plasmaconcentrations (rat) at day 3 (FIG. 2) and day 28 (FIG. 3).

DETAILED DESCRIPTION OF THE INVENTION

The methods for producing a lyophilized degarelix product both accordingto the first and the second aspect start with the degarelix drugsubstance which will be described in more detail.

The Degarelix Drug Substance

The decapeptide degarelix can be prepared by solid phase peptidesynthesis, as disclosed in WO98/46634, WO2010/12835 and WO2011/066386,or by liquid phase peptide synthesis, as disclosed in WO2012/055905 orWO2012/055903. This peptide synthesis provides crude degarelix which isfurther purified and then lyophilized to provide a lyophilized productwhich consists of degarelix, acetic acid, a residual amount of water,and minor amounts of impurities due to the production process, if any.This product is referred to as degarelix drug substance or merely drugsubstance in the present invention. The degarelix drug substancepreferably consists of degarelix, 4.5 to 10 wt. % acetic acid (w/w), andup to 10 wt. % water (w/w).

The manufacture of the drug substance can be divided into a first step(A) providing purified degarelix in solution, and a second step (B)providing the degarelix drug substance.

Step (A): Purification

Step (A) comprises the purification of crude degarelix in one or moresteps, preferably two steps, optionally followed by a columnconcentration and/or salt exchange step.

Crude degarelix, as obtained by LPPS or SPPS, is first subjected topurification. The purification is preferably carried out by applying thepeptide solution obtained by SPPS or LPPS to a column with reversedphase material, which is preferably pre-equilibrated with buffer. Thisfirst purification step preferably provides a purity of at least 95%, asdetermined by HPLC.

In a preferred embodiment, the reverse phase column chromatography isrepeated to obtain a product with purity of at least 97.5%, asdetermined by HPLC.

In a particularly preferred embodiment, the purified degarelix solutionwith a purity of at least 95%, preferably at least 97.5%, is subjectedto further column chromatography step to pre-concentrate the degarelixsolution and/or for salt exchange (particularly if acetic acid is usedfor pH adjustment of the eluents in the last purification step).

This pre-concentration and/or salt exchange step is also preferablycarried out on a reverse phase column: The purified degarelix solutionis diluted with water (preferably 1.5 to 2.5 times) and applied to thecolumn, pre-equilibrated with buffer. The column is preferably firstwashed with ethanol (low concentration, generally below 20%) and aqueousammonium acetate and subsequently with ethanol (low concentration,generally below 20%)/acetic acid/water. The column is then eluted, e.g.with ethanol (high concentration, generally 20 to 60%)/acetic acid/waterto obtain a more concentrated solution of degarelix compared to thesolutions after the purification step(s). The process is not limited toethanol as an organic modifier in the eluent. Other solvents such asacetonitrile can also be used.

Step (A) provides purified degarelix in solution.

Step (B)

The subsequent treatment of the purified degarelix in solution to obtainthe degarelix drug substance can be carried out in different ways. Inthe following, four preferred ways are illustrated (Steps (B1), (B2),(B3), and (B4)).

Step (B1): Concentration-De-aggregation-Lyophilization

The purified degarelix in solution is first subjected to a concentrationstep in which ethanol or another organic modifier such as acetonitrileis removed by evaporation. This step is preferably carried out with arotavap evaporator. The preferred maximum temperature during evaporationis 40° C. The resulting highly concentrated, viscous degarelix product(aggregated product that is usually in gel form) is then treated withacetic acid (de-aggregation step), preferably filtered, and lyophilized.

The de-aggregation step is important to control the viscosity of thedrug substance. Therefore, the de-aggregation step is preferably carriedout with one or more, most preferably all of the following conditions:

-   -   Final acetic acid concentration: 6-40%, preferably 15-35% (v/v)    -   Temperature 0 to 35° C., preferably 2 to 30° C., most preferably        5 to 15° C.    -   Degarelix concentration (free base) 5-35 g/l, preferably 10-20        g/I    -   Time 1-15 hours

Within these limits, suitable combinations of process parameters can befound by simple experimentation.

The lyophilization step is preferably carried out at an ice thickness of1.2 to 2.4 cm and a secondary drying time of 1 to 17 hours. Thesecondary drying temperature is usually around 20° C. (15 to 25° C.).

In a preferred embodiment, step (B1) thus provides a lyophilized drugsubstance that shows a viscosity of less than 3.2 mPas, preferablybetween 1.15 and 2 mPas (as determined upon dissolution in an amount of20 mg degarelix free base in 1 ml of water, containing 2.5% (w/V)mannitol). The method for measuring the viscosity is described in theexperimental section. A drug substance fulfilling this viscosityrequirement is further referred to as drug substance (1), whereas a drugsubstance not fulfilling this viscosity requirement is further referredto as drug substance (2). Drug substance (2) has a preferred viscosityof 3.2 to 15 mPas, upon dissolution in an amount of 20 mg in 1 ml water.Drug substance (2) may even be gel-like, in a condition of significantlymore than 3.2 mPas, even though its viscosity cannot be measuredprecisely.

In this respect, it is noted that “upon dissolution in an amount of 20mg in 1 ml of water” merely refers to the conditions for measuring theviscosity and does not mean that the drug substance is present in asolution of 20 mg/ml. Most preferably, the drug substance is present inlyophilized form.

The drug substance, in particular drug substance (1), is preferablyfurther characterized by an acetic acid content of 4.5 to 10.0% (w/w)and/or a water content of 10% or less (w/w). Additionally, the drugsubstance, in particular drug substance (1), preferably shows an opticaldensity of 0.10 AU or less (at a concentration of 20 mg degarelix freebase/ml in 2.5% mannitol (aq)). The method for measuring the opticaldensity is described in the experimental section.

Thus, the invention provides a method for producing the degarelix drugsubstance (1), comprising the steps of:

-   -   a. Purifying degarelix as obtained by liquid or solid phase        peptide synthesis to obtain a degarelix solution with a purity        of at least 95%;    -   b. Evaporating solvent to concentrate the degarelix solution to        obtain aggregated degarelix;    -   c. Deaggregating the aggregated degarelix with acetic acid; and    -   d. Lyophilizing the deaggregated degarelix to provide the        degarelix drug substance.

The invention further provides a method for modulating the viscosity ofdegarelix, such that following lyophilization and reconstitution withwater, the viscosity of a 20 mg/ml degarelix solution in 2.5% w/Vmannitol is no greater than 15 mPas, comprising:

-   -   treating aggregated degarelix with acetic acid; and    -   lyophilizing the mixture of degarelix and acetic acid.

In certain embodiments, the conditions for acetic acid addition andlyophilization are as described above. In particular embodiments, theacetic acid content following lyophilization is 4.5 to 10.0% (w/w).

Step (B2): Column Concentration—Lyophilization

The degarelix solution obtained in step A (preferably after a one- ortwo-step purification method, e.g. without pre-concentration/saltexchange) is loaded onto a chromatographic column, ion exchange isperformed and the column is rinsed (washed) with diluted acetic acid(about 1%). Degarelix is eluted from the column using aqueous aceticacid at a AcOH concentration of 20 to 50 wt. %, preferably 23 to 37 wt.%, preferably 23 to 27 wt. %, preferably 27 to 37 wt. %, preferably 33to 37 wt. %, preferably 35 wt. %, and subsequently optionally diluted toappropriate AcOH concentration, filtered. Then, the AcOH/water solutionof degarelix is lyophilized. The stationary phase in the column can beof different types. The stationary phase can contain functional groupslike hydrocarbons (aliphatic and aromatic), alcohols, nitriles, groupswith appropriate acid/base properties and ion-exchange groups but is notlimited to this type of groups. This process provides the drugsubstance, preferably drug substance (1).

Thus, the invention also provides a method for producing the degarelixdrug substance, comprising the steps of:

-   -   a. Purifying degarelix as obtained by liquid or solid phase        peptide synthesis to obtain a degarelix solution with a purity        of at least 95%;    -   b. Loading the degarelix solution onto a chromatographic column;    -   c. Eluting degarelix from the column with acetic acid to provide        eluted degarelix;    -   d. Lyophilizing the eluted degarelix to provide the degarelix        drug substance.        Step (B3) Isolation Via Lyophilization-Reconstitution in        AcOH/Water-Lyophilization

Purified degarelix in solution obtained after Step A is isolated vialyophilization; the resulting lyophilized product is dissolved at aconcentration between 10 and 20 g/L in 2% acetic acid, and lyophilizedagain to give degarelix drug substance (1).

Step (B4): Spray-Drying

Purified degarelix in solution (EtOH/water containing AcOH or ACN/watercontaining AcOH obtained after Step A, or as described in Step B1 exceptfor lyophilization, or B2 except for lyophilization) is isolated viaspray drying to give degarelix drug substance (1).

In a preferred embodiment, a purified degarelix solution as obtained instep A is directly subjected to the spray-drying process. The AcOHconcentration of said aqueous degarelix solution subjected tospray-drying is adjusted to 6 to 40% (v/v), preferably 15 to 35% (v/v).

After having described the manufacture of the drug substance, we willnow describe the manufacture of degarelix drug product. The lyophilizeddegarelix drug product comprises the degarelix drug substance andmannitol, i.e. it comprises (and preferably consists of) degarelix,acetic acid, mannitol, a residual amount of water, and minor amounts ofimpurities due to the production process, if any.

Production Process a of the Degarelix Drug Product

Production process A is the first aspect of the invention mentionedabove, i.e. a method for producing degarelix drug product which, uponreconstitution with water for injection in an amount of 20 mg degarelixfree base/ml, shows a viscosity of up to 15 mPas, preferably within arange of 2 to 12 mPas, comprising the steps of:

-   -   a. Providing a lyophilized degarelix drug substance, preferably        degarelix drug substance (1);    -   b. Dissolving the lyophilized degarelix drug substance in        mannitol-containing water to provide an aqueous        degarelix-mannitol mixture;    -   c. Lyophilizing the aqueous degarelix-mannitol mixture to        provide the degarelix drug product.

In the present invention, “upon reconstitution with water for injectionin an amount of 20 mg/ml” refers to the conditions for measuring theviscosity and does not mean that the drug product is present in asolution of 20 mg/ml. Most preferably, the drug product is present inlyophilized form, optionally in combination with reconstitution liquid.Preferred amounts per vial are in the range of 60 to 300 mg (such as 120mg, 80 mg, and 240 mg). Alternatively, it can be provided asreconstituted drug product, with preferred concentrations in the rangeof 2 to 100 mg/ml, preferably 10 to 70 mg/ml (such as 40 mg/ml, 20mg/ml, and 60 mg/ml).

Step b may also be referred to as compounding step. In a preferredembodiment, filtration and vial filling are carried out aftercompounding and before freeze-drying so that the entire preferredproduction process A comprises the steps of:

-   -   Compounding to provide the unfiltered bulk drug product    -   Filtration (sterile)    -   Vial filling    -   Freeze-drying/Lyophilization        Compounding to Provide the Bulk Drug Product

The drug substance is subjected to a compounding step, which isgenerally carried out as follows:

For the production of the unfiltered bulk drug product, drug substanceand mannitol are dissolved in water (pure water; generally Milli-Qwater) each in amounts of 10-60 g, per 1000 g batch size. Typicalamounts are 20 to 50 g drug substance (as degarelix free base content asdetermined by HPLC and 10 to 50 g mannitol per 1000 g. The actual amountdepends on the final concentration of degarelix in the drug product andthe volume of the reconstitution liquid (mannitol is preferably addedsuch that an isotonic solution with an osmolality of 300 mOsm+/−30 mOsmis obtained after reconstitution).

For production, water (usually approx. 80% of the total amount of water)is added to a compounding vessel. The mannitol is added and dissolved bystirring. Then the drug substance is added to the stirred mannitolsolution and the formulated bulk (batch) is brought to its final weightby adding the remaining water. This compounding is carried out in amanner so that a significant viscosity increase is avoided. Theviscosity of the bulk product thus preferably remains below 5 mPas,preferably below 3.2 mPas during the compounding step (viscositydetermined after filtration upon dissolution in an amount of 20 mg in 1ml of 2.5% (w/V) aqueous mannitol solution). This can be achieved bywetting the peptide at a high stirring speed for a relatively short timeperiod (up to 30 minutes) and then dissolving the peptide at a reducedstirring speed to avoid foaming and aggregation (generally for 30 to 90minutes). The temperature is usually kept within a range of 6-15° C. Thestirrer is preferably one that provides turbulent mixing without vortex.

Filtration

The bulk drug product is then preferably sterile filtered, e.g. throughtwo sterilizing grade filters placed in series, by pressurizing theformulated bulk with nitrogen.

Filling

Sterilized vials are filled with the filtered bulk drug product andsemi-stoppered (freeze-drying position) under aseptic conditions.

Freeze-Drying

The freeze-dryer is preferably steam sterilized before use. The vialsare the placed on the freeze-dryer shelves. The subsequent freeze-dryingprocess preferably comprises the steps of freezing, main drying(sublimation), and secondary drying. Preferred conditions are asfollows:

The freeze-drying process preferably comprises, or even consists of,three main steps, i.e. freezing, main drying (sublimation) and secondarydrying.

Freezing

The vials are loaded onto refrigerated shelves maintained at 2 to 10°C., such as 5° C.

The shelves are cooled from e.g. 5° C. to −30 to −40° C., such as −35°C. The shelf temperature is maintained at e.g. −35° C. for minimum twohours to ensure complete freezing of the entire batch prior tocommencement of primary drying.

Main Drying (Sublimation)

Main drying is performed by lowering the chamber pressure (preferably to0.100 mBar or less) and increasing the shelf temperature (preferably to10 to 20° C., such as +17° C.).

The main drying time proceeds for at least 15 hours.

Secondary Drying

After completion of the primary dying process, chamber pressure isreduced (preferably to 0.01 mBar or less) and the shelf temperature isincreased (preferably to 20 to 30° C., such as 25° C.). Secondary dryingis typically completed within 7 hours.

The lyophilized drug product is then labeled and packaged and combinedwith the appropriate amount of reconstitution liquid.

The reconstitution liquid is selected depending on the viscosity of thedrug substance. If drug substance (1) is used as starting material forproduction process A, i.e. a lyophilized drug substance that shows aviscosity of less than 3.2 mPas, preferably between 1.15 and 2 mPas(upon dissolution in an amount of 20 mg in 1 ml of 2.5 (w/v) % aqueousmannitol solution), the resulting lyophilized drug product is preferablycombined with water for injection (WFI) as reconstitution liquid. If thelyophilized drug product is reconstituted with WFI, the viscosity isgenerally in the range of 2 to 15 mPas (as measured upon dissolution of20 mg degarelix (free base) in 1 ml WFI). A drug product with aviscosity within this range was found to provide a sufficient depotrelease of degarelix in vivo.

Production Process B of the Degarelix Drug Product

Production process B is a method for producing degarelix drug productcomprising a lyophilized degarelix drug product and a liquid forreconstitution (reconstitution fluid) which, upon reconstitution withsaid liquid in an amount of 20 mg degarelix free base/ml, shows aviscosity of up to 15 mPas, preferably within a range of 2 to 12 mPas,comprising the steps of:

-   -   a. Dissolving a lyophilized degarelix drug substance in a        mannitol containing aqueous solution to provide a        degarelix-mannitol mixture;    -   b. Lyophilizing the degarelix-mannitol mixture to provide the        degarelix drug product,        wherein a viscosity-reducing agent is added to the        degarelix-mannitol mixture prior to lyophilization.

Step a may also be referred to as compounding step. In a preferredembodiment, filtration and vial filling are carried out aftercompounding and before freeze-drying so that the entire preferredproduction process B comprises the steps of:

-   -   Compounding to provide the unfiltered bulk drug product    -   Filtration (sterile)    -   Vial filling    -   Freeze-drying/Lyophilization

Production process B is identical to production process A, with theexception that in the compounding step, a viscosity-reducing agent,preferably a non-ionic surfactant is added prior to lyophilization. Thenon-ionic surfactant is preferably added in an amount of 0.0003 to 1.5mg/ml to the bulk solution, corresponding to an amount of 0.001 to 5mg/ml, more preferably 0.1 to 1 mg/ml, in the reconstituted drug product(e.g. when reconstituted to a degarelix concentration of 60 mg degarelixfree base/ml). Preferred non-ionic surfactants are those with a linearalkyl chain having at least 8 carbon atoms (preferably without doublebonds) and a carbohydrate moiety. Particularly preferred are those thatare approved for subcutaneous injections, such as Tween 20 (Polysorbate20). Other suitable non-ionic surfactants includeTocopheryl-polyethylene-glycol-1000-succinate (TPGS) and other Tweens.

As starting material for production process B, drug substance (2) ispreferably used, i.e. a lyophilized drug substance that shows aviscosity of at least 3.2 mPas (upon dissolution in an amount of 20 mgdegarelix free base in 1 ml of 2.5 wt. % aqueous mannitol solution). Theresulting lyophilized drug product is typically combined with WFI asreconstitution liquid. If the lyophilized drug product is reconstitutedwith WFI, the viscosity is generally in the range of 2 to 15 mPas (asmeasured upon dissolution of 20 mg degarelix free base in 1 ml WFI). Adrug product with a viscosity within this range was found to provide asufficient depot formation for delayed release of degarelix in vivo.

Production process B is particularly preferred for degarelix productsthat have a relatively high degarelix concentration upon reconstitution,such as 50 mg degarelix free base/ml or more, e.g. 60 mg degarelix freebase/ml (240 mg drug product).

Novel Degarelix Drug Product

Production process B provides a novel degarelix drug product thatdiffers from known drug products in that the reconstituted drug productcontains a viscosity-reducing agent. The viscosity-reducing agent is theone used in step B, preferably a non-ionic surfactant.

The present invention thus provides a degarelix drug product whichcomprising a lyophilized degarelix drug product and a liquid forreconstitution which, upon reconstitution with said liquid in an amountof 20 mg degarelix free base/ml, shows a viscosity of up to 15 mPas,preferably within a range of 2 to 12 mPas, and contains aviscosity-reducing agent in an amount of 0.001 to 5 mg/ml, morepreferably 0.1 to 1 mg/ml.

EXPERIMENTAL SECTION Example 1: Purification, Deaggregation andLyophilisation

Crude Degarelix was synthesized as described in WO2012/055905 A1, up toStep 12 in Example 5. Step 13 as disclosed in WO2012/055905 A1 wasreplaced by the following steps, In summary, the purification process ofcrude degarelix drug substance, obtained after the last deprotectionstep, consists of three preparative reversed phase chromatography (RPC)steps, where the third RPC step primarily is a desalting step.

Step 13 (Purification Step):

Crude degarelix solution from step 12 in WO02012/055905A1 was applied toa column packed with reversed phase material, pre-equilibrated withbuffer (90% of 0.12% TFA and 10% of EtOH). Load: ≤30 g/L column volume.The column was washed and eluted with a gradient: Buffer (EtOH 29% to 50and 0.12% TFA (aq) 71% to 50%). The fractions obtained were analysed,and combined in such a way that the purity of the main pool fulfilledthe acceptance criterion for the process control.

When elution was checked by HPLC method, purity is ≥95%.

Step 14 (Purification Step):

The main pool obtained in the step 13 was diluted twice with water andapplied to the column packed with reversed phase material,pre-equilibrated with buffer (90% of 1% AcOH and 10% of EtOH). Load: ≤25g/L column volume. The column was first washed with a first buffer (10%of EtOH and 90% of 0.5 mol/L AcONH4) and then with a second buffer (90%of 1% AcOH and 10% of EtOH).

The column was then eluted with a mixture of buffer and ethanol (76% of1% AcOH and 24% of EtOH). The fractions obtained were analyzed, andcombined in such a way that the purity in the main pool fulfilled theacceptance criteria for the process control.

When elution was checked by “HPLC method”, purity is ≥97.5%.

Step 15 (Pre-Concentration/Salt Exchange):

The main pool obtained in the step 14 was diluted twice with water andapplied to the column packed with reversed phase material,pre-equilibrated with buffer (90% of 1% AcOH and 10% of EtOH). Load: ≤18g/L column volume. The column was first washed with a first buffer (10%of EtOH and 90% of 0.5 mol/L AcONH4) and then with a second buffer (90%of 1% AcOH and 10% of EtOH). The column was then eluted with a mixtureof buffer and ethanol (50% of 1% AcOH and 50% of EtOH).

As an alternative, degarelix can be eluted with a solution ofAcOH/MeCN/water, such as 12% AcOH and 22% MeCN in water.

Step 16 (Concentration-De-Aggregation-Lyophilization):

Prior to lyophilisation, the pool of pure degarelix solutions from step15 was concentrated below 40° C. Aqueous acetic acid and water wereadded to the concentrated solution, to give a concentration below 15g/L, and an acetic acid concentration of 30%. This solution was thenfiltered and lyophilised to yield degarelix drug substance.

Pressure: ≤0.5 mbar during the primary and secondary drying

Temperatures: End freezing temperature: ≤−30° C.

-   -   End primary drying temperature: 20° C.    -   End secondary drying temperature: 20° C.        Time: 5 days        Results

The viscosity of the drug substance obtained by the sequence of stepswas below 2.5 mPas, as determined in a concentration of 20 mg/ml in 2.5(w/V) % mannitol solution.

Example 2: Method for Measuring Viscosity for Drug Substance and DrugProduct

Determination of the viscosity of the drug substance and drug productsolution is based on the current edition of Ph. Eur. and USP procedureusing a rotating viscometer equipped with a cone-plate measuring systemwith D=60 mm and 1°. The shear rate is increased from 0 to 500 s⁻¹ in 20steps, using a controlled rate (CR) rotation step program at a constanttemperature of 20±0.2° C., making certain that the system reachesequilibrium before the viscosity is recorded at a shear rate of 500 s⁻¹.

Example 3: Method for Measuring Optical Density for Drug Substance andDrug Product

Equipment and Materials

-   -   UV spectrophotometer    -   UV light transmitting cuvettes, 10 mm path.    -   Cuvette lids    -   Water for Injection (used for reconstitution of degarelix drug        product and in the reference cell)        Performance of Analysis

Between reconstitution and measurement the reconstituted samples must bekept at 22° C.±1° C.

The drug substance vial is reconstituted 2.5% aqueous mannitol solutionin water (w/V). The drug product vial is reconstituted with water forinjection. Dispense the solvent into the vial and swirl the vial untilreconstitution is complete or use a vortex for a few seconds. The liquidshould look clear and no undissolved powder or particles are visible.Keep the vial upright and do not shake. The sample is measured at 350nm, 120 minutes after addition of the solvent.

Four minutes before the measurement, the sample has to be homogenized bygently turning the cuvette five times back and forth throughapproximately 180 degrees. The four minute delay allows any air bubblesto disperse before the reading.

The absorption caused by the cuvette and by the water for injection hasto be deducted from the read-off of the sample.

Example 4: Spray Drying

Preparation of Feed

In the preparation of the feed, the weighed amount of degarelix, shownin the table below, was dissolved using a magnetic stirrer. Thedifferent feed solutions, W—VI and AI-IV were prepared by addition ofMilli-Q water, or pure glacial acetic acid (99.9%) in the followingmanner:

-   a. Batch W-IV: the appropriate volume of water was added to the    weighed amount of degarelix-   b. Batch A-I to A-Ill: for each experiment, acetic acid solutions    were prepared by diluting 99.0% glacial acetic acid with Milli-Q    water resulting in 30, 5, 2 percent solutions, to which the weighed    amount of degarelix as added

Prior to all of the spray-drying runs, the reconstituted peptide wasfiltered into a measuring flask through a 0.20 μm Sartoriuos Ministarfilter prior to spray-drying.

Spray-Drying

Prior to spray-drying, the inlet temperature and the feed rate wereadjusted. The tubing of the pump was placed in the feed solution, andthe drying was initiated. When the drying was completed, the inlettemperature was allowed to drop to 700° C. before the cyclone and thecollection vessel were dismantled for powder collection. The powder wascollected with brushes into Petri dishes, which were weighed before andafter collection to determine the yield.

Overview of the Setting Applied for Spray-Drying

Parameters W-IV A-I A-II A-III Inlet temperature, 120 120 120 120 ° C.Outlet 85 85 85 85 temperature, ° C. Atomising air 600 600 600 600 flow,L/h Liquid feed rate, 3 3 3 3 mL/min. Aspirator rate, 30 30 30 30 m3/hSolvent water Aq. 30% Aq. 5% Aq. 2% AcOH AcOH AcOH Solution volume, 2525 25 25 mL Amount of 1 1 1 1 degarelix Concentration of 4 4 4 4 feed, %

The viscosity of reconstituted degarelix spray-dried from acetic acidsolutions of four different concentrations (A-I to A-Ill) compared toone batch of degarelix spray-dried from water (W-IV)

Batch no. W-IV A-I A-II A-III Viscosity, 2.40 1.68 2.11 2.10 mPas * S

Example 5: Column Concentration—Lyophilization

The pool from step 14 of Example 1 was diluted with water and applied toa column packed with reversed phase material. After rinsing the columnwith 1% AcOH in water degarelix was eluted with 35% AcOH in water.Fractions were adjusted to contain 35 g degarelix/l, 27% AcOH (Sample 1)and 15 g degarelix/l, 30% AcOH (Sample 2). The adjusted fractions werefreeze-dried and analysed. Results: see Table below:

Analysis on drug substance Sample 1 Sample 2 Water (w/w) 2.1% 2.5%Residual AcOH (w/w) 7.2% 7.0% OD 20 mg/ml (15 min/6 h) <0.02 AU/0.02 AU<0.02 AU/0.02 AU Viscosity 20 mg/ml 1.88 mPas 1.94 mPas

Example 6: Manufacture of Degarelix Drug Substance and DeaggregationStudies

Materials

De-aggregation studies of purified, crude drug substance

-   -   Crude, purified degarelix was supplied at a concentration of        57.4 mg/ml    -   Acetic acid 100% was supplied by Merck    -   De-aggregation equipment:        -   Compounding vessel: 300 ml double walled glass vessel        -   Blue Cap bottles, 50 or 100 ml        -   Magnet        -   Magnetic stirrer    -   Primary packaging:        -   Colourless 20R glass vials. Vials were washed and dried in a            heat chamber.        -   20 mm freeze-drying stoppers type I according to Ph. Eur/USP            20 mm Flip-Off Caps

Manufacturing of Drug Products Using the De-Aggregated Drug Substances

-   -   Two experimental drug substances were used. As a control, a        commercial drug substance is used.    -   Mannitol: D (−) mannitol (PF-05-0232)    -   Primary packaging:        -   Colourless 10R glass vials (according to DIN ISO 8362).            Vials are washed and dried in a heat chamber.        -   20 mm freeze-drying stoppers type I according to Ph. Eur/USP            (type 1319, rubber W4416/50/grey, The West Company)        -   20 mm Flip-Off Caps (The West Company)

Methods

De-Aggregation Experiments Using a Drug Substance Stock Solution

Temperature During the Experiments:

During the de-aggregation studies the temperature was set to 5, 20, 25,or 35° C.

Concentration of Degarelix Drug Substance in Stock Solutions During theExperiments:

The concentration of degarelix drug substance was 5, 15, 25, or 35mg/ml.

Concentration of Acetic Acid During the Experiments:

The concentration of acetic acid is 13, 15, 18, 20, 22, 26, 30, 35 or40%.

Analysis Performed During the Experiments:

Viscosity: Take out ˜1.2 ml at T=1, 60, 120, and 240 min.

If it fits better into the schedule to take the sample at another timepoint, then this can be done provided the correct time is written intothe formulation record. However, the sampling point at T=1 min should bekept.

Optical Density (OD): A sample (˜1 ml) should be taken at the end ofeach experiment for measurement of the final optical density.

De-aggregation Experiment Protocol, total volume 20 or 69 ml:

-   -   1. Weigh the required amount of stock solution into a beaker.        Equilibrate to the desired temperature    -   2. Weigh the required amount of acetic acid (100%) into a beaker    -   3. Mix the acetic acid (100%) with 3 or 10 ml of milli-Q water        and equilibrate to the desired temperature    -   4. Mix the degarelix stock solution with the acetic acid/milli-Q        water solution    -   5. Fill milli-Q water up to 20 or 69 ml    -   6. Mix thoroughly    -   7. Take samples for viscosity and optical density as described        above. Note the appearance of the solution during the experiment        Settings for De-Aggregation of the Two 69 ml Degarelix Stock        Solution:        Degarelix drug substance conc. 25 mg/ml in both experiments        Acetic acid: 10% or 13%        Temperature: 5° C.

Filling the two test de-aggregated drug substances (69 ml) into vialsand freeze-drying: The de-aggregated solutions were filled immediatelyinto 20R vials. 5 ml were filled into each vial. The vials and bulkshould be kept cold during filling (preferably between 5-10° C.). Assoon as the vials were filled they were placed in the freeze dried andthe program was initiated.

Subsequent to freeze drying and closing of the vials, the vials werestored in the freeze dryer at 5° C. until de-loading.

The two freeze dried drug substances were subjected to the followinganalyses:

Viscosity was measured on samples of 20 mg/ml free base

Optical density is measured on samples of 20 mg/ml

Content of degarelix

Content of acetate

Results

AcOH Viscosity Temp. Degarelix conc. conc. @ 500 mPa * s Optical Density(° C.) mg/ml (%) Time = 240 min Time = 240 min 5 5 40 1.9 0.00 5 25 132.3 0.10 5 25 15 1.9 0.09 5 25 20 1.7 0.06 5 35 26 1.9 0.08 20 15 30 1.80.03 25 25 20 2.0 0.10 35 25 15 7.2 0.86 35 25 18 2.1 0.07 35 25 20 1.70.05 35 35 22 2.1 0.13 35 35 26 1.9 0.10 35 35 30 2.0 0.07 35 35 35 2.10.08

Example 7: Production of Drug Product without Viscosity-Reducing Agent

Compounding of Lab-Scale Drug Product Batches Using the TwoDe-Aggregated Drug Substances

The excipients listed below in Table 1 are used in all compoundingexperiments.

Excipients Mannitol Milli-Q waterCompounding Equipment:Compounding vessel: 300 ml double walled glass vesselBlue Cap bottles, 100 mlMagnetMagnetic stirrerPrimary Packaging:

-   -   Colourless 10R glass vials. Vials were washed and dried in a        heat chamber.    -   20 mm freeze-drying stoppers type I    -   20 mm Flip-Off Caps        Study Description

Degarelix bulk solutions containing 20 mg/g degarelix and 25 mg/gmannitol are compounded using different batches of drug substance anddifferent settings of temperature.

The viscosities of the bulk solutions is measured during/aftercompounding on two occasions (after dissolution and at t=120 minutes).

Bulk Composition:

The batch size and composition of the two bulk solutions is presented inTable 2.

TABLE 1 Batch size and composition of the bulk solutions Batch size (g)50 Drug substance (g) 1.000 g/degarelix free base content drug substanceMannitol (g)  1.25 Milli-Q water - Total (g) Ad. 50Experimental Design:

The compounding parameters are given below in Table 2.

TABLE 2 Compounding parameters Time Parameters Drug substance Test drugsubstance batches Temperature 15° C. Stirring speed 50 rpm Water,starting volume 80%Compounding:

The compounding is performed with the stirrer positioned at the centre.Before commencing the experiments the centre position should be fixed.

-   -   1. Weigh the required ingredients        -   40 g of Milli-Q water into the compounding vessel.        -   1.25 g of mannitol into a suitable container.        -   Weigh exact amount of degarelix drug substance (see Table 2)            into a suitable container.        -   Weigh the remaining amount of Milli-Q water into a suitable            container.    -   2. Connect the double walled compounding vessel containing the        Milli-Q water to the cooling circulator. Connect a second double        walled vessel in series. The container containing the rest water        is placed in the second double walled vessel in order that it        will equilibrate to the correct temperature before being added        to the bulk (step 9).    -   3. Carefully place the stirring magnet in the compounding        vessel. Set the stirring rate to 50 rpm using the tachometer.    -   4. Set the cooler to the correct temperature, start the stirring        and add the mannitol to the compounding vessel. Stir until        mannitol has dissolved.    -   5. Allow the system to equilibrate to the set temperature.    -   6. Measure the temperature in the mannitol solution.    -   7. Remove the stirrer. Start the timer and immediately add the        drug substance. Then reinsert the stirrer.    -   8. At t=5 minutes add the rest of the Milli-Q water, evenly over        the surface of the drug substance using a glass Pasteur pipette.    -   9. When only a few lumps are left, rinse down drug substance        from the sides of the vessel with the dissolution medium.    -   10. When the drug substance is completely dissolved make a note        of the time and measure the temperature. Take out a sample for        viscosity measurement of the bulk.    -   11. Continue the experiment until t=120 minutes. Take out a        sample for viscosity measurement of the bulk.        Filtering

The bulks will not be filtered because it is expected that at least oneof the batches will be highly aggregated and therefore very difficult tofilter. Also, the batches will not be used for purposes where sterilityis required.

Filling

Fill 6.40 g bulk into 10R vials. The vials and bulk should be kept coldduring filling (preferably between 5-10° C.). As soon as the vials arefilled they should be placed in the freeze dried and the program shouldbe initiated. Each bulk should result in approx. 6-7 vials.

Results

18 different drug substance batches, with different viscosities asindicated in the table below, were processed as described above. Theviscosity of the corresponding drug products was determined as follows:

Viscosity DS (20 mg/ml) Viscosity DP (20 mg/ml) Batch mPas mPas 1 2.35.8 2 2.3 7.81 3 2.1 4.75 4 2.3 8.62 5 2.1 7.85 6 2.1 7.09 7 2.3 7.76 82.1 7.69 9 2.1 7.96 10 2.3 8.1 11 1.8 7.82 12 1.7 4.14 13 2.3 8.46 142.3 9.57 15 2.3 10.96 16 2.3 8.47 17 2.5 8.05 18 2.5 10.4

These results are graphically illustrated in FIG. 1. FIG. 1 shows acorrelation between drug substance viscosity and drug product viscosity.

Example 8: Production of Drug Product with Viscosity-Reducing AgentExperiment 1: Surfactants Tested (Reconstitution Using WFI/SurfactantSolution)

Procedure:

-   -   Degarelix batch reconstituted using WFI (as control) and using        WFI containing the surfactants. The vials are reconstituted to        60 mg/ml degarelix. The drug product batch used for these        experiments had a bulk viscosity of approx. 3.8 mPas.    -   TPGS and Tween 20 were prepared in solutions containing 1 mg/ml        (0.1% solutions)    -   OD (absorbance) was measured at 350 nm    -   Absorbance was registered after 30 min, 60 min, 120 min and 24        hours Result:        Both TPCG and Tween 20 reduced the optical density to a level        below the control.

Experiment 2: Reconstitution of Degarelix Vials with [WFI+Tween 20] doesnot Alter the In-Vitro Release Profile (IVD Assay) at 40 mg/ml ofDegarelix

Tween 20 concentrations tested:

1 mg/ml; 0.5 mg/ml; 0.25 mg/ml;

Control: WFI

Results: The in-vitro release profiles are not altered by the additionof Tween-20.

Example 9: Drug Product Viscosity and Bioavailability

Summary

A multivariate data analysis was undertaken with a set of data from n=38batches of degarelix drug products, including in vivo pharmacokineticdata (rat model).

A relationship between physico-chemical characteristics and in vivopharmacokinetics in a rat model was established. The study revealed thatthe viscosity of the reconstituted degarelix product appears to be theprominent and only parameter with some ability to predict in vivoperformance of the depot.

Introduction

Degarelix drug product is manufactured as a freeze-dried productcontaining mannitol. The products are used as investigational medicinalproducts in clinical studies. Several formulations were produced,containing various amounts of degarelix per vial, namely 10 mg, 20 mg,40 mg, 88 mg, 128 mg, 120 mg (40 mg/ml), 180 mg (60 mg/ml) and 240 mg(60 mg/ml) and different ratios of degarelix/mannitol.

Materials

Approximately 40 different batches of degarelix have been produced withdifferent batches of degarelix substance and formulations. The standarddegarelix concentration in the bulk is 20 g/I unless otherwise stated.

Methods

Physico-Chemical Methods

The different methods to characterise aggregation were selected frommultivariate data analysis. Data from n=38 batches were compiled.However, smaller populations were used for some methods that were onlyimplemented with some formulations (e.g. measurements at 40 mg/ml notperformed with the 20 mg formulation).

Rat Bioassay

A standardised rat assay consisting in following the pharmacokineticprofile of degarelix over 28 days was used. In order to avoid local sideeffects, the rats were given degarelix as a 20 mg/ml suspension with aninjection volume of 100 μl. Groups of n=8 rats were used, all treatedwith the reconstituted suspension originating from one single vial.Plasma concentrations were initially measured at 2 hrs, 1 day, 7 day and28 day and partial AUC calculated as AUC 0-7 days and AUC 7-28 days. Thedesign was then changed with measurement of plasma concentration (Cp)-2hrs being skipped and being replaced by Cp-3 days. Likewise, AUC 0-7days was replaced by AUC 1-7 days. This explains the discontinuities inthe population sizes of some of the in vivo variables.

Multivariate Data Analysis

Given the size of the data set, an approach by multivariate dataanalysis (PCA and PLS) with the Simca-P, version 10 software (UmetricsAB, SE-Umeå) was implemented. Data were handled as previously, includingsoft block scaling (¼ root), scaling to unit variance (UV) andcentering. Turbidity data were log transformed to improve datadistribution. In vivo Cp data were also log transformed to stabilise thevariance.

Relationship Between Drug Product Physico-Chemistry and In VivoPerformance in a Rat Model

Relationship Between Physico-Chemistry and In Vivo Performance

A first model was calculated based on 2 components. The goodness of fit(R2=0.58) was relatively low given the high variability of thebiological data (20-30%), but the goodness of predictability (Q2=0.35)was satisfactory (R2-Q2 should be in the range of 0.2 to 0.3). It couldbe seen from the loading scatter plot (not shown) that variables such asturbidity measured at 20 mg/ml and acetate content were not influential.

A new model was therefore generated excluding these 3 variables (2turbidity variables measured at 20 mg/ml and acetate), based on 2significant components and yielding to similar data description(R2=0.53) but improved predictability (Q2=0.42). Best explained andpredicted rat variables were plasma concentrations at 3 days and 28 days(Cp-3d, Cp-28d), and the area under the curve between day 1 and day 7(AUC 1-7d).

The coefficient overview plot indicated that larger influences werecarried by the viscosity data respectively at 20 mg/ml and then at 40mg/ml, all significant at a confidence level of 0.95 for everybiological variable. Specific surface area and turbidity were clearly oflesser influence and were not significant, even at a confidence level of0.90.

The relationship between dynamic viscosity measured at 20 mg/ml and bestfitted biological variables is shown in FIGS. 2 and 3.

CONCLUSION

A solid relationship (R2=0.53) was established between viscosity of theconstituted product and in vivo performance as investigated in a ratmodel. In both cases, a higher viscosity yielded to a reduced releasefrom the depot. Other physico-chemical variables were not relevant withthe exception of acetate content for in vitro dissolution. Therefore,the viscosity of constituted degarelix product appears to be theprominent parameter with some ability to predict in vitro release and invivo performance of the depot.

The invention claimed is:
 1. A lyophilized degarelix drug substancehaving a viscosity of up to 3.2 mPas, as determined upon dissolution inan amount of 20 mg degarelix free base in 1 ml of water containing 2.5%(w/V) mannitol, and the lyophilized degarelix drug substance has anacetic acid concentration ranging from 4.5% to 10.00% (w/w) and a watercontent of 10% (w/w) or less, wherein the lyophilized degarelix drugsubstance is prepared by a method, comprising: a. purifying degarelixobtained by a liquid or solid phase peptide synthesis to obtain adegarelix solution with a purity of at least 95%, as determined byhigh-performance liquid chromatography (HPLC); b. evaporating a solventto concentrate the degarelix solution to obtain aggregated degarelix; c.deaggregating the aggregated degarelix with acetic acid to obtaindeaggregated degarelix; and d. lyophilizing the deaggregated degarelixto provide the lyophilized degarelix drug substance.
 2. The substanceaccording to claim 1, wherein the purity of the degarelix solutionobtained in step (a) is at least 97.5%, as determined by HPLC.
 3. Thesubstance according to claim 1, wherein step (a) further comprises apre-concentration step comprising pre-equilibrating a pre-concentrationcolumn with a pre-equilibrating buffer; diluting the degarelix withwater; loading the degarelix onto the pre-concentration column; washingthe pre-concentration column with a first washing buffer comprisingethanol and aqueous ammonium acetate, and then a second washing solutioncomprising ethanol, acetic acid, and water; and eluting degarelix fromthe pre-concentration column.
 4. The substance according to claim 1,wherein the evaporating of step (b) is carried out at or below 40° C. 5.The substance according to claim 1, wherein the deaggreagated degarelixobtained in step (c) has an acetic acid concentration ranging from 6% to40% (v/v).
 6. The substance according to claim 1, wherein step (c) iscarried out over 1 to 15 hours.
 7. The substance according to claim 1,wherein step (d) comprises a primary drying procedure and a secondarydrying procedure, both of which are conducted at a pressure less than orequal to 0.5 mbar.
 8. The substance according to claim 7, wherein theprimary drying procedure is carried out at a primary drying temperatureof about 20° C.
 9. The substance according to claim 7, wherein thesecondary drying procedure is carried out at a secondary dryingtemperature ranging from 15° C. to 25° C.
 10. The substance according toclaim 1, wherein step (d) is carried out over a period of about 5 days.11. A method of preparing a lyophilized degarelix drug substance,comprising: a. purifying degarelix as obtained by liquid or solid phasepeptide synthesis to obtain a degarelix solution with a purity of atleast 95%, as determined by high-performance liquid chromatography(HPLC); b. isolating the degarelix solution via lyophilization to obtainisolated degarelix; c. dissolving the isolated degarelix to obtain areconstituted degarelix solution, wherein the reconstituted degarelixsolution of step (c) has a degarelix concentration ranging from 10 to 20g degarelix/L of aqueous 2% acetic acid; and d. lyophilizing thereconstituted degarelix solution to provide the lyophilized degarelixdrug substance, wherein the lyophilized degarelix drug substance has aviscosity of less than 3.2 mPas, as determined upon dissolution in 1 mLof aqueous 2.5% (w/V) mannitol solution of an amount of the lyophilizeddegarelix drug substance necessary to achieve a degarelix concentrationof 20 mg/mL; and wherein the lyophilized degarelix drug substance has anacetic acid concentration ranging from 4.5% to 10.00% (w/w) and a watercontent of 10% (w/w) or less.
 12. The method according to claim 11,wherein the purity of the degarelix solution obtained in step (a) is atleast 97.5%, as determined by HPLC.
 13. A method of preparing adegarelix drug substance, comprising the following steps a, b, and d; ora, c, and d: a. purifying degarelix as obtained by liquid or solid phasepeptide synthesis to obtain a degarelix solution with a purity of atleast 95%, as determined by high-performance liquid chromatography(HPLC); and b. evaporating a solvent to concentrate the degarelixsolution to obtain aggregated degarelix and deaggregating the aggregateddegarelix with acetic acid; c. loading the degarelix solution onto achromatographic column and eluting degarelix from the column with aceticacid; d. isolating the degarelix via spray drying to provide thedegarelix drug substance, and wherein the degarelix drug substance has aviscosity of less than 3.2 mPas, as determined upon dissolution in 1 mLof aqueous 2.5% (w/V) mannitol solution of an amount of the degarelixdrug substance necessary to achieve a degarelix concentration of 20mg/mL; and wherein the degarelix drug substance has an acetic acidconcentration ranging from 4.5% to 10.00% (w/w) and a water content of10% (w/w) or less.
 14. The method according to claim 13, wherein thepurity of the degarelix solution obtained in step (a) is at least 97.5%,as determined by HPLC.
 15. The method according to claim 13, wherein thespray drying of step (d) comprises an inlet temperature of about 120°C., an outlet temperature of about 85° C., an atomizing air flow rate ofabout 600 L/h, a liquid feed rate of about 3 mL/min, and an aspiratorrate of about 30 m³/h.
 16. A method of preparing a degarelix drugsubstance, comprising: a. purifying degarelix as obtained by liquid orsolid phase peptide synthesis to obtain a degarelix solution with apurity of at least 95%, as determined by high-performance liquidchromatography (HPLC); and b. isolating the degarelix via spray dryingto provide the degarelix drug substance, wherein the degarelix drugsubstance provided has a viscosity of less than 3.2 mPas, as determinedupon dissolution in 1 mL of aqueous 2.5% (w/V) mannitol solution of anamount of the degarelix drug substance necessary to achieve a degarelixconcentration of 20 mg/mL; and wherein said degarelix drug substance hasan acetic acid concentration ranging from 4.5% to 10.00% (w/w) and awater content of 10% (w/w) or less.
 17. A lyophilized degarelix drugsubstance prepared by the method according to claim
 11. 18. Alyophilized degarelix drug substance prepared by the method according toclaim
 13. 19. A lyophilized degarelix drug substance prepared by themethod according to claim 16.